Abstract:
:A method for use in high-throughput screening of bacteria for the production of long-chain hydrocarbons and ketones by monitoring fluorescent light emission in the presence of Nile red is described. Nile red has previously been used to screen for polyhydroxybutyrate (PHB) and fatty acid esters, but this is the first report of screening for recombinant bacteria making hydrocarbons or ketones. The microtiter plate assay was evaluated using wild-type and recombinant strains of Shewanella oneidensis and Escherichia coli expressing the enzyme OleA, previously shown to initiate hydrocarbon biosynthesis. The strains expressing exogenous Stenotrophomonas maltophilia oleA, with increased levels of ketone production as determined by gas chromatography-mass spectrometry, were distinguished with Nile red fluorescence. Confocal microscopy images of S. oneidensis oleA-expressing strains stained with Nile red were consistent with a membrane localization of the ketones. This differed from Nile red staining of bacterial PHB or algal lipid droplets that showed intracellular inclusion bodies. These results demonstrated the applicability of Nile red in a high-throughput technique for the detection of bacterial hydrocarbons and ketones.
journal_name
mBiojournal_title
mBioauthors
Pinzon NM,Aukema KG,Gralnick JA,Wackett LPdoi
10.1128/mBio.00109-11subject
Has Abstractpub_date
2011-06-28 00:00:00pages
e00109-11issue
4issn
2150-7511pii
mBio.00109-11journal_volume
2pub_type
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